Primary biliary cirrhosis (PBC) is an autoimmune hepatic disorder characterized by the immune mediated destruction of bile ducts and the presence of high titer antimitochondrial antibodies. PBC is a frequently lethal disease and patients with PBC account for 30-40% of all adult liver transplant recipients in the USA, The autoimmune response in PBC is directed to enzymes of the 2-oxo dehydrogenase family and there is extensive data on autoantibody reactivity to these proteins. The major aims of this grant are to elucidate the nature of the T cell mediated response and to determine why immune damage is confined to the biliary epithelium despite the ubiquitous distribution of the target autoantigens. Key reagents previously developed or used in prior work including cDNAs encoding autoantigens, panels of well-defined sera, availability of liver biopsies, liver from transplant recipients, hepatic cDNA libraries from patients with PBC, biliary epithelial cultures and T cell cloned lines specific for mitochondrial autoantigens will be used. In the T cell work, T cell cloned lines will be derived from patients with PBC, other liver diseases and normals. The T cell cloned lines will be examined for phenotype and T cell receptor usage by a combination of FACS sorting, hybridization with specific gene probes such as T cell receptor genes or lymphokines. T cell receptor sequence will be determined by nucleotide sequencing. The epitopes that T cell recognize will be mapped by a combination of recombinant autoantigen fragments, tryptic digests of native proteins and synthetic peptides. Questions of autoantigen production will be investigated using specific antisera and biliary cell cultures by immunofluorescence and immunochemical techniques. The work on autoantibodies and epitope mapping will be continued for those antigens where data is lacking. An attempt will be made to correlate precise measurements of antibody specificity, titer, affinity and subclass with clinical or prognostic indicators. We believe that the definition of the autoimmune response to the mitochondrial autoantigens at the molecular level will further our understanding and may lead to specific protocols for the treatment of PBC.